Facioscapulohumeral muscular dystrophy (FSHD) is the third most common form of muscular dystrophy and affects 1 in 15,000 to 20,000 people worldwide. Developing and testing therapeutics for FSHD would be significantly advanced if a valid mouse model of the disease were available. Ideally, a murine version of FSHD would reproduce all the features of FSHD muscle, retaining the morphological, physiological and genomic differences found in fresh biopsies. As the pathophysiological mechanisms that lead to FSHD are still unclear, it would be best if such a model were constructed from FSHD tissue itself, rather than by manipulating particular genes or gene products that, although involved, may not be sufficient for pathogenesis. Finally, the mice should be accessible to many laboratories and so should be available in significant numbers. My colleagues and I have been developing such a model. In particular, we have been investigating the use of xenografting to create mice with humanized Tibialis anterior muscles developed from FSHD and control myogenic cell lines. In our preliminary studies, we have used methods that are routine in our laboratory, including X-irradiation of the lower hindlimbs of immunocompromised (NRG) mice and injection of cardiotoxin to eliminate the Tibialis anterior muscles of the mice and their abiliy to regenerate. We have also incorporated new methods, including periodic electrical stimulation of the graft through the peroneal nerve, and unique reagents and methods provided by our collaborators, Drs. W.E. Wright (UT Southwestern), P. Jones (UMass Med School) and L.M. Kunkel (Boston Children's Hospital). Our preliminary results, obtained with a human myogenic cell line that expresses luciferase, to facilitate monitoring by luminometry, show that these cells engraft into the otherwise empty TA compartment of mice, where they survive to form mature muscle fibers that are human in origin and that are essentially free of murine myonuclei. The human fibers are innervated and they contract upon stimulation of the peroneal nerve, to generate a specific force comparable to that of human muscle. As the grafts contain PAX7-positive human cells located adjacent to human myofibers, they are also likely to have the capacity for further growth. Recent data with a cell line derived from an FSHD patient indicate that these cells engraft efficiently as well, forming muscles that specifically express Dux4 and two of its downstream targets. This suggests that our methods preserve the genetic characteristics of myogenic cells as they engraft, grow, and differentiate. We have two specific aims: (i) to optimize the formation of mature human muscle tissue in mice, as a first step in developing a model of FSHD for therapeutic testing; and (ii) to apply our optimized methods to cells from FSHD patients and their unaffected relatives, to develop, characterize, and assess the variability of a novel model of FSHD in mice, for subsequent therapeutic testing. Our studies should therefore generate a model of FSHD muscle in mice that will be an invaluable reagent in identifying and testing drugs to treat FSHD.